Characterization studies have been carried out on a protein kinase which is stimulated by both calcium and phospholipids. Differentiated PYS endoderm cells possess calcium, phospholipid - dependent protein kinase (Ca, PL-PK) activity while no such activity could be detected in undifferentiated F9 cells. Retinoic acid- induced differentiation of F9 cells to an endoderm cell type provoked a time-dependent increase in cytosolic Ca, PL-PK activity. This increase in Ca, PL-PK activity correlates with diofferentiation to the parietal endoderm cell type. Employing EL 4 thymoma cells as a source of enzyme, it was found that cytosolic Ca, PL-PK activity is markedly decreased following treatment of the cells with phorbol ester. The decrease occurs within 5 min. after the addition of phorbol ester and is dose dependent. Alteration of this protein kinase activity conceivably mediates later effects of the phorbol ester tumor promoter. It was found that the calcium binding protein, calmodulin, contains a calcium-induced hydrophobic site. Using hydrophobic interaction chromatography a single step procedure was developed to rapidly purify calmodulin to homogeniety.